This is a competitive renewal of an NCI grant to study the contributions of trisomy 21 to hematologic malignancies. Children with DS, who have trisomy 21 in all their cells, are remarkably predisposed to leukemia, with an estimated 1 in 10 newborns exhibiting transient myeloproliferative disorder (TMD) and 1 in 500 DS children developing acute megakaryocytic leukemia (AMKL) by the age of five. In addition to myeloid leukemia, children with DS have a 20-fold increased risk of B-cell acute lymphoblastic leukemia (B-ALL). We hypothesize that overexpression of a small number of genes in the Down syndrome critical region directly lead to increased incidence of leukemia. With a greater knowledge of these genes on human chromosome 21 (Hsa21) and the specific events that occur in the evolution of these diseases, improved diagnostics and therapies can be discovered. During the previous funding period, we made many important discoveries regarding the role of trisomy 21 and GATA1 mutations in these diseases, including 1) the demonstration that mouse models of DS develop myeloproliferative disease characterized by aberrant megakaryopoiesis, 2) a detailed characterization of the role of GATA1 mutations on megakaryocyte growth and development, 3) the discovery of mutations in signaling pathways in the evolution of AMKL, 4) creation of a faithful animal model of DS-AMKL, and 6) the identification of the genes ERG and DYRK1A as the most likely leukemia oncogenes on human chromosome 21. In this renewal, we will use a combination of animal models and human patient samples to precisely determine the contributions of trisomy 21 to AMKL and ALL. Specifically, we will: 1) Determine the necessity and sufficiency of selected Hsa21 genes in human AMKL and the murine model of DS-AMKL, 2) Identify the signaling pathways that are activated in AMKL, such as NFAT which lies downstream of DYRK1A, and 3) Determine the contribution of trisomy 21 to B-ALL. In addition to providing insights into the role of trisomy 21 in DS leukemia, this research is also relevant to other leukemias with acquired trisomy 21, such as hyperdiploid ALL and to myeloproliferative neoplasms, in particular the large group of ET and PMF cases that lack JAK2 or MPL mutations.